The histone deacetylases inhibitors (HDACi) are a promising new class of anti-cancer drugs. Their utility in the clinic is currently under intensive research. HDACi cause acetylation of histones and thereby weaken the tight bond between DNA and histones. This relaxes the DNA and may render it more accessible to molecules that interact with DNA, such as topoisomerase II (topo II) inhibitors. Topo II inhibitors are widely used in clinical practice in several cancer types; however they are limited by resistance and by their narrow therapeutic margins. Our in vitro data indicate that combining HDACi in a specific schedule with topo II inhibitors is synergistic. Furthermore, this synergy is seen predominantly in cells with "normal" or increased levels of topo lIalpha and Ilbeta and is associated with histone hyperacetylation and a paradoxical decrease in expression of topo IIalpha, but not IIbeta. We found that several HDACi synergized with topo II inhibitors in preclinical assays. The anti-seizure drug valproic acid (VPA), which is known to be an HDACi showed potent synergy in several cancer cell lines. An advantage of using this drug is its well-understood and comprehensively documented toxicity and safety profile. In preclinical studies, this synergy was independent of cell origin but dependant on pretreatment topo II expression. In Specific Aim 1, we propose to conduct a Phase I trial combining the HDACi, VPA and the topo II inhibitor epirubicin in a specific sequence evaluating toxicity and tolerability of this combination. In Specific Aims 2 and 3, we will collect and analyze pharmacological and molecular endpoints of this trial to determine whether there is A) an association between VPA and epirubicin drug levels with toxicity and outcome, B) a dose response relationship between VPA concentrations and histone hyperacetylation, C) a difference in VPA-induced histone hyperacetylation in peripheral blood mononuclear cells and tumors cells and D) an association between pre-treatment expression levels and VPA-induced changes of topo Ilalpha and IIbeta in tumor samples with response. This proposal may help to establish a new paradigm for sequence-specific combinations of HDACi and topo inhibitors and to define a methodology to study predictive and surrogate markers of response. These findings may help to establish VPA as a clinically useful HDACi and to direct the rational design of future clinical trials involving other HDACi. Finally, the utility of the topo II inhibitors could be enhanced if their efficacy could be increased without worsening toxicity, as our preclinical data appears to indicate.